The vascular gene Apold1 is dispensable for normal development but controls angiogenesis under pathological conditions.

The molecular mechanisms of angiogenesis have been intensely studied, but many genes that control endothelial behavior and fate still need to be described. Here, we characterize the role of Apold1 (Apolipoprotein L domain containing 1) in angiogenesis in vivo and in vitro. Single-cell analyses reveal that - across tissues - the expression of Apold1 is restricted to the vasculature and that Apold1 expression in endothelial cells (ECs) is highly sensitive to environmental factors. Using Apold1-/- mice, we find that Apold1 is dispensable for development and does not affect postnatal retinal angiogenesis nor alters the vascular network in adult brain and muscle. However, when exposed to ischemic conditions following photothrombotic stroke as well as femoral artery ligation, Apold1-/- mice display dramatic impairments in recovery and revascularization. We also find that human tumor endothelial cells express strikingly higher levels of Apold1 and that Apold1 deletion in mice stunts the growth of subcutaneous B16 melanoma tumors, which have smaller and poorly perfused vessels. Mechanistically, Apold1 is activated in ECs upon growth factor stimulation as well as in hypoxia, and Apold1 intrinsically controls EC proliferation but not migration. Our data demonstrate that Apold1 is a key regulator of angiogenesis in pathological settings, whereas it does not affect developmental angiogenesis, thus making it a promising candidate for clinical investigation.

Early life stress affects the miRNA cargo of epididymal extracellular vesicles in mouse†.

Sperm RNA can be modified by environmental factors and has been implicated in communicating signals about changes in a father's environment to the offspring. The small RNA composition of sperm could be changed during its final stage of maturation in the epididymis by extracellular vesicles (EVs) released by epididymal cells. We studied the effect of exposure to stress in early postnatal life on the transcriptome of epididymal EVs using a mouse model of transgenerational transmission. We found that the small RNA signature of epididymal EVs, particularly miRNAs, is altered in adult males exposed to postnatal stress. In some cases, these miRNA changes correlate with differences in the expression of their target genes in sperm and zygotes generated from that sperm. These results suggest that stressful experiences in early life can have persistent biological effects on the male reproductive tract that may in part be responsible for the transmission of the effects of exposure to the offspring.

Effects of Early Life Stress on Bone Homeostasis in Mice and Humans.

Bone pathology is frequent in stressed individuals. A comprehensive examination of mechanisms linking life stress, depression and disturbed bone homeostasis is missing. In this translational study, mice exposed to early life stress (MSUS) were examined for bone microarchitecture (µCT), metabolism (qPCR/ELISA), and neuronal stress mediator expression (qPCR) and compared with a sample of depressive patients with or without early life stress by analyzing bone mineral density (BMD) (DXA) and metabolic changes in serum (osteocalcin, PINP, CTX-I). MSUS mice showed a significant decrease in NGF, NPYR1, VIPR1 and TACR1 expression, higher innervation density in bone, and increased serum levels of CTX-I, suggesting a milieu in favor of catabolic bone turnover. MSUS mice had a significantly lower body weight compared to control mice, and this caused minor effects on bone microarchitecture. Depressive patients with experiences of childhood neglect also showed a catabolic pattern. A significant reduction in BMD was observed in depressive patients with childhood abuse and stressful life events during childhood. Therefore, future studies on prevention and treatment strategies for both mental and bone disease should consider early life stress as a risk factor for bone pathologies.

Single-Cell Multiomics Techniques: From Conception to Applications.

Recent advances in methods for single-cell analyses and barcoding strategies have led to considerable progress in research. The development of multiplexed assays offers the possibility to conduct parallel analyses of multiple factors and processes for comprehensive characterization of cellular and molecular states in health and disease. These technologies have expanded extremely rapidly in the past years and constantly evolve and provide better specificity, precision and resolution. This review summarizes recent progress in single-cell multiomics approaches, and focuses, in particular, on the most innovative techniques that integrate genome, epigenome and transcriptome profiling. It describes the methodologies, discusses their advantages and limitations, and explains how they have been applied to studies on cell heterogeneity and differentiation, and epigenetic reprogramming.

High Efficiency RNA Extraction From Sperm Cells Using Guanidinium Thiocyanate Supplemented With Tris(2-Carboxyethyl)Phosphine.

The extraction of high-quality ribonucleic acid (RNA) from tissues and cells is a key step in many biological assays. Guanidinium thiocyanate-phenol-chloroform (AGPC) is a widely used and efficient method to obtain pure RNA from most tissues and cells. However, it is not efficient with some cells like sperm cells because they are resistant to chaotropic lysis solutions containing guanidinium thiocyanate such as Buffer RLT+ and Trizol. Here, we show that disulfide bonds are responsible for the chemical resistance of sperm cells to RNA extraction reagents. We show that while ß-mercaptoethanol (ßME) can increase sperm lysis in Buffer RLT+, it has no effect in Trizol and leaves sperm cells intact. We measured the reduction of disulfide bonds in 2,2'-dithiodipyridine (DTDP) and observed that ßME has a pH-dependent activity in chaotropic solutions, suggesting that pH is a limiting factor. We identified tris(2-carboxyethyl)phosphine (TCEP) as an efficient lysis enhancer of AGPC solutions that can retain reducing activity even at acidic pH. Trizol supplemented with TCEP allows the complete and rapid lysis of sperm cells, increasing RNA yield by 100-fold and resulting in RNA with optimal quality for reverse transcription and polymerase chain reaction. Our findings highlight the importance of efficient cell lysis and extraction of various macromolecules for bulk and single-cell assays, and can be applied to other lysis-resistant cells and vesicles, thereby optimizing the amount of required starting material and animals.

Symposium summary: Epigenetic inheritance-impact for biology and society 26-28 August 2019, Zurich, Switzerland.

The concept of epigenetic inheritance proposes a new and unconventional way to think about heredity in health and disease, at the interface between genetics and the environment. Epigenetic inheritance is a form of biological inheritance not encoded in the DNA sequence itself but mediated by epigenetic factors. Because epigenetic factors can be modulated by the environment, they can relay this information to the genome and modify its activity consequentially. If epigenetic changes induced by environmental exposure are present in the germline and persist in germ cells during development until conception, they have the potential to transfer the traces of ancestral exposure to the progeny. This form of heredity relates to the extremely important question of nature versus nurture and how much of our own make-up is genetically or epigenetically determined, a question that remains largely unresolved. Because it questions the dominant dogma of genetics and brings a paradigm shift in sciences, it has to creating strong bridges between disciplines and provide solid causal evidence to be firmly established. The second edition of a conference fully dedicated to epigenetic inheritance was held in August 2019 in Zurich, Switzerland. This symposium titled 'Epigenetic inheritance: impact for biology and society' (http://www.epigenetic-inheritance-zurich.ethz.ch), gathered experts in the field of epigenetic inheritance to discuss the concept and pertinent findings, exchange views and expertise about models and methods, and address challenges raised by this new discipline. The symposium offered a mix of invited lectures and short talks selected from abstracts, poster sessions and a workshop 'Meet the experts: Q&A'. A tour of a local omics facility the Functional Genomics Center Zurich was also offered to interested participants. Additional comments and impressions were shared by attendees on Twitter #eisz19 during and after the symposium. This summary provides an overview of the different sessions and talks and describes the main findings presented.

Transgenerational Epigenetics of Traumatic Stress.

Traumatic stress is a type of environmental experience that can modify behavior, cognition and physiological functions such as metabolism, in mammals. Many of the effects of traumatic stress can be transmitted to subsequent generations even when individuals from these generations are not exposed to any traumatic stressor. This book chapter discusses the concept of epigenetic/non-genomic inheritance of such traits involving the germline in mammals. It includes a comprehensive review of animal and human studies on inter- and transgenerational inheritance of the effects of traumatic stress, some of the epigenetic changes in the germline currently known to be associated with traumatic stress, and possible mechanisms for their induction and maintenance during development and adulthood. We also describe some experimental interventions that attempted to prevent the transmission of these effects, and consider the evolutionary importance of transgenerational inheritance and future outlook of the field.

Transgenerational inheritance of behavioral and metabolic effects of paternal exposure to traumatic stress in early postnatal life: evidence in the 4th generation.

In the past decades, evidence supporting the transmission of acquired traits across generations has reshaped the field of genetics and the understanding of disease susceptibility. In humans, pioneer studies showed that exposure to famine, endocrine disruptors or trauma can affect descendants, and has led to a paradigm shift in thinking about heredity. Studies in humans have however been limited by the low number of successive generations, the different conditions that can be examined, and the lack of mechanistic insight they can provide. Animal models have been instrumental to circumvent these limitations and allowed studies on the mechanisms of inheritance of environmentally induced traits across generations in controlled and reproducible settings. However, most models available today are only intergenerational and do not demonstrate transmission beyond the direct offspring of exposed individuals. Here, we report transgenerational transmission of behavioral and metabolic phenotypes up to the 4th generation in a mouse model of paternal postnatal trauma (MSUS). Based on large animal numbers (up to 124 per group) from several independent breedings conducted 10 years apart by different experimenters, we show that depressive-like behaviors are transmitted to the offspring until the third generation, and risk-taking and glucose dysregulation until the fourth generation via males. The symptoms are consistent and reproducible, and persist with similar severity across generations. These results provide strong evidence that adverse conditions in early postnatal life can have transgenerational effects, and highlight the validity of MSUS as a solid model of transgenerational epigenetic inheritance.

Stress does not increase blood-brain barrier permeability in mice.

Several studies have reported that exposure to acute psychophysiological stressors can lead to an increase in blood-brain barrier permeability, but these findings remain controversial and disputed. We thoroughly examined this issue by assessing the effect of several well-established paradigms of acute stress and chronic stress on blood-brain barrier permeability in several brain areas of adult mice. Using cerebral extraction ratio for the small molecule tracer sodium fluorescein (NaF, 376 Da) as a sensitive measure of blood-brain barrier permeability, we find that neither acute swim nor restraint stress lead to increased cerebral extraction ratio. Daily 6-h restraint stress for 21 days, a model for the severe detrimental impact of chronic stress on brain function, also does not alter cerebral extraction ratio. In contrast, we find that cold forced swim and cold restraint stress both lead to a transient, pronounced decrease of cerebral extraction ratio in hippocampus and cortex, suggesting that body temperature can be an important confounding factor in studies of blood-brain barrier permeability. To additionally assess if stress could change blood-brain barrier permeability for macromolecules, we measured cerebral extraction ratio for fluorescein isothiocyanate-dextran (70 kDa). We find that neither acute restraint nor cold swim stress affected blood-brain barrier permeability for macromolecules, thus corroborating our findings that various stressors do not increase blood-brain barrier permeability.

Rapid stress-induced transcriptomic changes in the brain depend on beta-adrenergic signaling.

Acute exposure to stressful experiences can rapidly increase anxiety and cause neuropsychiatric disorders. The effects of stress result in part from the release of neurotransmitters and hormones, which regulate gene expression in different brain regions. The fast neuroendocrine response to stress is largely mediated by norepinephrine (NE) and corticotropin releasing hormone (CRH), followed by a slower and more sustained release of corticosterone. While corticosterone is an important regulator of gene expression, it is not clear which stress-signals contribute to the rapid regulation of gene expression observed immediately after stress exposure. Here, we demonstrate in mice that 45 min after an acute swim stress challenge, large changes in gene expression occur across the transcriptome in the hippocampus, a region sensitive to the effects of stress. We identify multiple candidate genes that are rapidly and transiently altered in both males and females. Using a pharmacological approach, we show that most of these rapidly induced genes are regulated by NE through ß-adrenergic receptor signaling. We find that CRH and corticosterone can also contribute to rapid changes in gene expression, although these effects appear to be restricted to fewer genes. These results newly reveal a widespread impact of NE on the transcriptome and identify novel genes associated with stress and adrenergic signaling.

Hippocampal gene expression induced by cold swim stress depends on sex and handling.

Stress-related disorders such as PTSD and depression are more prevalent in women than men. One reason for such discordance may be that brain regions involved in stress responses are more sensitive to stress in females. Here, we compared the effects of acute stress on gene transcription in the hippocampus of female and male mice, and also examined the involvement of two key stress-related hormones, corticosterone and corticotropin releasing hormone (Crh). Using quantitative reverse transcription polymerase chain reaction (RT-qPCR), we measured gene expression of Fos, Per1 and Sgk1 45 min after exposure to brief cold swim stress. Stress induced a stronger increase in Fos and Per1 expression in females than males. The handling control procedure increased Fos in both sexes, but occluded the effects of stress in males. Further, handling increased Per1 only in males. Sgk1 was insensitive to handling, and increased in response to stress similarly in males and females. The transcriptional changes observed after swim stress were not mimicked by corticosterone injections, and the stress-induced increase in Fos, Per1 and Sgk1 could neither be prevented by pharmacologically blocking glucocorticoid receptor (GR) nor by blocking Crh receptor 1 (Crhr1) before stress exposure. Finally, we demonstrate that the effects are stressor-specific, as the expression of target genes could not be increased by brief restraint stress in either sex. In summary, we find strong effects of acute swim stress on hippocampal gene expression, complex interactions between handling and sex, and a remarkably unique response pattern for each gene. Overall, females respond to a cold swim challenge with stronger hippocampal gene transcription than males, independent of two classic mediators of the stress response, corticosterone and Crh. These findings may have important implications for understanding the higher vulnerability of women to certain stress-related neuropsychiatric diseases.

[Surgical treatment of tumors of the central region of brain in children: analysis of own material and review of the literature]. / Leczenie operacyjne nowotworów obszaru srodkowego mózgu u dzieci--analiza materialu wlasnego i przeglad pismiennictwa.

UNLABELLED: AIM OF THIS PAPER: Is to present our results of surgical treatment of central region tumors in children and to determine prognostic factors for patients' survival. METHOD: Retrospective analysis of medical records, radiographies and histopathological specimens of patients treated at the Dept. of Neurosurgery with the diagnosis of central region tumor and review of questionnaires mailed to the patients' parents. The statistical analysis was performed using the "Statistica.pl" software. MATERIAL: From January 1980 to March 2003, 114 such children were treated (mean age--8.05 years, mean follow-up time--38.4 months). In the subgroup of benign tumors (n=70), 70% of children are still alive 21.4% are dead and no data are available for the remaining 8.5% (mean survival time--23 months). In the subgroup of malignant tumors (n=12), 33.3% of patients are still alive and 66.6% are dead (mean survival time--14.2 months). In the group treated surgically (total or partial resection) (n=35), 71.4% of children are still alive, 17.1% are dead and no data are available in 11.4% of cases (mean survival time--24 months). In the group of children who underwent palliative procedures only (n=47), 59.5% are still alive, 27% are dead and no data are available for the remaining 12.7% (mean survival time--18.3 months). The statistical analysis revealed highly significant differences in the survival of children dependent on the tumor grade (p=0.00059) and the extent of resection (p=0.00825). CONCLUSIONS: Tumors of the central region of brain in children are highly heterogenous, so in every case the histological diagnosis should be obtained. Prognostic factors are the degree of histologic malignancy and the extent of surgical resection. In every case surgical resection should be considered.